Despite advances in model systems for identifying genes involved in aging, many unanswered questions still remain about the biology and underlying mechanisms in humans, particularly related to age-related cognitive decline. This is, in part, because even between mouse and human genomes, genetic regulatory elements have diverged, in sequence, numbers, and their presence in different genes. Additionally, the extreme heterogeneity of cell types in brain is a factor. In this grant, we take on the challenging problem of genetic mechanisms underlying human brain aging. First, we exploit the accumulated wealth of knowledge of the transcriptional repressor, REST, to fully characterize its function in neurons in adult brain. We, and others, have been studying REST for decades as a model for understanding fundamentals of gene regulation because of some prominent features. REST has an easily recognizable unusually large binding site that occurs in thousands of neuronal genes and it robustly signals its function when bound to genes by recruiting chromatin modifiers. While others have reported an activator function for REST in neurons, we have only found repressor function, so we will also settle this discussion in Aim 1. Aim 1 also includes studies to identify ?orphan? REST binding sites of unknown function. The results from this aim will expand the neuronal functions under REST control, as well as provide a potentially new model of repressor function, generally. Second, our preliminary results indicate REST is expressed to different levels in different neuronal types, human and mouse, and that its loss in mice results in differential transcriptional effects in these neurons. This parsing of REST function in human neurons is studied in depth in Aims 2 and 3. We recently found that REST protein levels in human hippocampus increase with age sometime in the second decade after birth, remaining elevated even in centenarians, whereas REST levels in mouse brain are greatly reduced by 4-6m after birth, and stay extremely low for life, suggesting human-specific REST functions and/or target genes with age. We will identify REST functions with age in Aim 2. Third, polymorphisms in REST have been associated with the age-related disease, Alzheimer's Disease (AD). A recent study suggests that REST is involved directly in cognitive decline, although some of the published findings are at odds with our results. We will test hypotheses regarding REST gene regulation in AD in Aim 3. In total, our proposed experiments will provide new information on REST function and its potential impact on neuronal gene expression and chromatin changes during brain aging and related pathologies. The experiments will also provide new information on fundamental questions regarding repressor function.